Thrombopoietin mimetics

ABSTRACT

Invented are non-peptide TPO mimetics. Also invented are novel processes and intermediates used in the preparation of the presently invented compounds. Also invented is a method of treating thrombocytopenia, in a mammal, including a human, in need thereof which comprises administering to such mammal an effective amount of a selected hydroxy-1-azobenzene derivative.

This application is a continuation of U.S. application Ser. No.11/650,651, filed Jan. 8, 2007, now U.S. Pat. No. 7,473,686 which is acontinuation of U.S. application Ser. No. 10/296,688, filed Jul. 3,2003, now U.S. Pat. No. 7,160,870, granted Jan. 9, 2007, which is a 371of International Application No. PCT/US01/16863, filed May 24, 2001,which claims the benefit of U.S. Provisional Application No. 60/228,929,filed Aug. 30, 2000 and U.S. Provisional Application No. 60/207,084,filed May 25, 2000.

FIELD OF THE INVENTION

This invention relates to thrombopoietin (TPO) mimetics and their use aspromoters of thrombopoiesis and megakaryocytopoiesis.

BACKGROUND OF THE INVENTION

Megakaryocytes are bone marrow-derived cells, which are responsible forproducing circulating blood platelets. Although comprising <0.25% of thebone marrow cells in most species, they have >10 times the volume oftypical marrow cells. See Kuter et al. Proc. Natl. Acad. Aci. USA 91:11104-11108 (1994). Megakaryocytes undergo a process known asendomitosis whereby they replicate their nuclei but fail to undergo celldivision and thereby give rise to polypoid cells. In response to adecreased platelet count, the endomitotic rate increases, higher ploidymegakaryocytes are formed, and the number of megakaryocytes may increaseup to 3-fold. See Harker J. Clin. Invest. 47: 458-465 (1968). Incontrast, in response to an elevated platelet count, the endomitoticrate decreases, lower ploidy megakaryocytes are formed, and the numberof megakaryocytes may decrease by 50%.

The exact physiological feedback mechanism by which the mass ofcirculating platelets regulates the endomitrotic rate and number of bonemarrow megakaryocytes is not known. The circulating thrombopoieticfactor involved in mediating this feedback loop is now thought to bethrombopoietin (TPO). More specifically, TPO has been shown to be themain humoral regulator in situations involving thrombocytopenia. See,e.g., Metcalf Nature 369:519-520 (1994). TPO has been shown in severalstudies to increase platelet counts, increase platelet size, andincrease isotope incorporation into platelets of recipient animals.Specifically, TPO is thought to affect megakaryocytopoiesis in severalways: (1) it produces increases in megakaryocyte size and number; (2) itproduces an increase in DNA content, in the form of polyploidy, inmegakaryocytes; (3) it increases megakaryocyte endomitosis; (4) itproduces increased maturation of megakaryocytes; and (5) it produces anincrease in the percentage of precursor cells, in the form of smallacetylcholinesterase-positive cells, in the bone marrow.

Because platelets (thrombocytes) are necessary for blood clotting andwhen their numbers are very low a patient is at risk of death fromcatastrophic hemorrhage, TPO has potential useful application in boththe diagnosis and the treatment of various hematological disorders, forexample, diseases primarily due to platelet defects (see Harker et al.Blood 91: 4427-4433 (1998)). Ongoing clinical trials with TPO haveindicated that TPO can be administered safely to patients (see Basser etal. Blood 89: 3118-3128 (1997); Fanucchi et al. New Engl. J. Med. 336:404-409 (1997)). In addition, recent studies have provided a basis forthe projection of efficacy of TPO therapy in the treatment ofthrombocytopenia, and particularly thrombocytopenia resulting fromchemotherapy, radiation therapy, or bone marrow transplantation astreatment for cancer or lymphoma. (See Harker, Curr. Opin. Hematol. 6:127-134 (1999)).

The gene encoding TPO has been cloned and characterized. See Kuter etal., Proc. Natl. Acad. Sci. USA 91: 11104-11108 (1994); Barley et al.,Cell 77: 1117-1124 (1994); Kaushansky et al., Nature 369:568-571 (1994);Wendling et al., Nature 369: 571-574 (1994); and Sauvage et al., Nature369: 533-538 (1994). Thrombopoietin is a glycoprotein with at least twoforms, with apparent molecular masses of 25 kDa and 31 kDa, with acommon N-terminal amino acid; sequence. See, Baatout, Haemostasis 27:1-8 (1997); Kaushansky, New Engl. J. Med. 339: 746-754 (1998).Thrombopoietin appears to have two distinct regions separated by apotential Arg-Arg cleavage site. The amino-terminal region is highlyconserved in man and mouse, and has some homology with erythropoietinand interferon-a and interferon-b. The carboxy-terminal region showswide species divergence.

The DNA sequences and encoded peptide sequences for human TPO receptor(TPO-R; also known as c-mpl) have been described. (See, Vigon et al.Proc. Natl. Acad. Sci. USA 89: 5640-5644 (1992)). TPO-R is a member ofthe haematopoietin growth factor receptor family, a family characterizedby a common structural design of the extracellular domain, including forconserved C residues in the N-terminal portion and a WSXWS motif closeto the transmembrane region. (See Bazan Proc. Natl. Acad. Sci. USA 87:6934-6938 (1990)). Evidence that this receptor plays a functional rolein hematopoiesis includes observations that its expression if restrictedto spleen, bone marrow, or fetal liver in mice (see Souyri et al. Cell63: 1137-1147 (1990)) and to megakaryocytes, platelets, and CD34+ cellsin humans (see Methia et al. Blood 82: 1395-1401 (1993)). Furtherevidence for TPO-R as a key regulator of megakaryopoiesis is the factthat exposure of CD34⁺ cells to synthetic oligonucleotides antisense toTPO-R RNA significantly inhibits the appearance of megakaryocytecolonies without affecting erythroid or myeloid colony formation. Someworkers postulate that the receptor functions as a homodimer, similar tothe situation with the receptors for G-CSF and erythropoietin. (seeAlexander et al. EMBO J. 14: 5569-5578 (1995)).

The slow recovery of platelet levels in patients suffering fromthrombocytopenia is a serious problem, and has lent urgency to thesearch for a blood growth factor agonist able to accelerate plateletregeneration (see Kuter, Seminars in Hematology, 37: Supp 4: 41-49(2000)).

It would be desirable to provide compounds which allow for the treatmentof thrombocytopenia by acting as a TPO mimetic.

As disclosed herein it has unexpectedly been discovered that certainhydroxy-1-azo-benzene derivatives are effective as agonists of the TPOreceptor, they are potent TPO mimetics.

SUMMARY OF THE INVENTION

This invention relates to compounds of Formula (I):

wherein:

-   -   R, R¹, R² and R³ are each independently selected from hydrogen,        C₁₋₆alkyl, —(CH₂)_(p)OR⁴, —C(O)OR⁴, formyl, nitro, cyano,        halogen, aryl, substituted aryl, substituted alkyl, —S(O)_(n)R⁴,        cycloalkyl, —NR⁵R⁶, protected —OH, —CONR⁵R⁶, phosphonic acid,        sulfonic acid, phosphinic acid, —SO₂NR⁵R⁶, and a heterocyclic        methylene substituent as represented by Formula (III),

-   -   -   where,        -   p is 0-6,        -   n is 0-2,        -   V, W, X and Z are each independently selected from O, S and            NR¹⁶, where R¹⁶ is selected from: hydrogen, alkyl,            cycloalkyl, C₁-C₁₂aryl, substituted alkyl, substituted            cycloalkyl and substituted C₁-C₁₂aryl,        -   R⁴ is selected from: hydrogen, alkyl, cycloalkyl,            C₁-C₁₂aryl, substituted alkyl, substituted cycloalkyl and            substituted C₁-C₁₂aryl, and        -   R⁵ and R⁶ are each independently selected from hydrogen,            alkyl, substituted alkyl, C₃₋₆cycloalkyl, and aryl,        -   or R⁵ and R⁶ taken together with the nitrogen to which they            are attached represent a 5 to 6 member saturated ring            containing up to one other heteroatom selected from oxygen            and nitrogen;

    -   m is 0-6; and

    -   AR is a cyclic or polycyclic aromatic ring containing from 3 to        16 carbon atoms and optionally containing one or more        heteroatoms, provided that when the number of carbon atoms is 3        the aromatic ring contains at least two heteroatoms and when the        number of carbon atoms is 4 the aromatic ring contains at least        one heteroatom, and optionally substituted with one or more        substituents selected from the group consisting of: alkyl,        substituted alkyl, aryl, substituted cycloalkyl, substituted        aryl, aryloxy, oxo, hydroxy, alkoxy, cycloalkyl, acyloxy, amino,        N-acylamino, nitro, cyano, halogen, —C(O)OR⁴, —C(O)NR¹⁰R¹¹,        —S(O)₂NR¹⁰R¹¹, —S(O)_(n)R⁴ and protected —OH,        -   where n is 0-2,        -   R⁴ is hydrogen, alkyl, cycloalkyl, C₁-C₁₂aryl, substituted            alkyl, substituted cycloalkyl and substituted C₁-C₁₂aryl,            and        -   R¹⁰ and R¹¹ are independently hydrogen, cycloalkyl,            C₁-C₁₂aryl, substituted cycloalkyl, substituted C₁-C₁₂aryl,            alkyl or alkyl substituted with one or more substituents            selected from the group consisting of: alkoxy, acyloxy,            aryloxy, amino, N-acylamino, oxo, hydroxy, —C(O)OR⁴,            —S(O)_(n)R⁴, —C(O)NR⁴R⁴, —S(O)₂NR⁴R⁴, nitro, cyano,            cycloalkyl, substituted cycloalkyl, halogen, aryl,            substituted aryl and protected —OH,        -   or R¹⁰ and R¹¹ taken together with the nitrogen to which            they are attached represent a 5 to 6 member saturated ring            containing up to one other heteroatom selected from oxygen            and nitrogen,        -   where R⁴ is as described above and n is 0-2;

    -   and pharmaceutically acceptable salts, hydrates, solvates and        esters thereof;

    -   provided that at least one of R, R¹, R² and R³ is a substituted        aryl group or a heterocyclic methylene substituent as        represented in Formula (III).

This invention relates to a method of treating thrombocytopenia, whichcomprises administering to a subject in need thereof an effective amountof a TPO mimetic compound of Formula (I).

The present invention also relates to the discovery that the compoundsof Formula (I) are active as agonists of the TPO receptor.

In a further aspect of the invention there is provided novel processesand novel intermediates useful in preparing the presently invented TPOmimetic compounds.

Included in the present invention are pharmaceutical compositionscomprising a pharmaceutical carrier and compounds useful in the methodsof the invention.

Also included in the present invention are methods of co-administeringthe presently invented TPO mimetic compounds with further activeingredients.

DETAILED DESCRIPTION OF THE INVENTION

This invention relates to compounds of Formula (I) as described above.

Included among the presently invented compounds of Formula (I) are thosehaving Formula (V):

wherein:

-   -   R, R¹, R² and R³ are each independently selected from hydrogen,        C₁₋₆alkyl, C₁₋₆alkoxy, —(CH₂)_(p)OR⁴, —C(O)OR⁴, formyl, nitro,        cyano, halogen, aryl, substituted aryl, substituted alkyl,        —S(O)_(n)R⁴, cycloalkyl, —NR⁵R⁶, protected —OH, —CONR⁵R⁶,        phosphonic acid, sulfonic acid, phosphinic acid and —SO₂NR⁵R⁶,        -   where,        -   p is 0-6,        -   n is 0-2,        -   R⁴ is selected from: hydrogen, alkyl, cycloalkyl,            C₁-C₁₂aryl, substituted alkyl, substituted cycloalkyl and            substituted C₁-C₁₂aryl, and        -   R⁵ and R⁶ are each independently selected from hydrogen,            alkyl, substituted alkyl, C₃₋₆cycloalkyl, and aryl,        -   or R⁵ and R⁶ taken together with the nitrogen to which they            are attached represent a 5 to 6 member saturated ring            containing up to one other heteroatom selected from oxygen            and nitrogen;    -   m is 0-6; and    -   AR is a cyclic or polycyclic aromatic ring containing from 3 to        16 carbon atoms and optionally containing one or more        heteroatoms, provided that when the number of carbon atoms is 3        the aromatic ring contains at least two heteroatoms and when the        number of carbon atoms is 4 the aromatic ring contains at least        one heteroatom, and optionally substituted with one or more        substituents selected from the group consisting of: alkyl,        substituted alkyl, aryl, substituted cycloalkyl, substituted        aryl, aryloxy, oxo, hydroxy, alkoxy, cycloalkyl, acyloxy, amino,        N-acylamino, nitro, cyano, halogen, —C(O)OR⁴, —C(O)NR¹⁰R¹¹,        —S(O)₂NR¹⁰R¹¹, —S(O)_(n)R⁴ and protected —OH,        -   where n is 0-2,        -   R⁴ is hydrogen, alkyl, cycloalkyl, C₁-C₁₂aryl, substituted            alkyl, substituted cycloalkyl and substituted C₁-C₁₂aryl;            and        -   R¹⁰ and R¹¹ are independently hydrogen, cycloalkyl,            C₁-C₁₂aryl, substituted cycloalkyl, substituted C₁-C₁₂aryl,            alkyl or alkyl substituted with one or more substituents            selected from the group consisting of: alkoxy, acyloxy,            aryloxy, amino, N-acylamino, oxo, hydroxy, —C(O)OR⁴,            —S(O)_(n)R⁴, —C(O)NR⁴R⁴, —S(O)₂NR⁴R⁴, nitro, cyano,            cycloalkyl, substituted cycloalkyl, halogen, aryl,            substituted aryl and protected —OH,        -   or R¹⁰ and R¹¹ taken together with the nitrogen to which            they are attached represent a 5 to 6 member saturated ring            containing up to one other heteroatom selected from oxygen            and nitrogen,        -   where R⁴ is as described above and n is 0-2;    -   and pharmaceutically acceptable salts, hydrates, solvates and        esters thereof;    -   provided that at least one of R, R¹, R² and R³ is a substituted        aryl group.

Preferred among the presently invented compounds of Formula (I) arethose having Formula (II):

wherein:

-   -   R, R¹, R² and R³ are each independently selected from hydrogen,        C₁₋₆alkyl, —(CH₂)_(p)OR⁴, —C(O)OR⁴, formyl, nitro, cyano,        halogen, aryl, substituted aryl, substituted alkyl, —S(O)_(n)R⁴,        cycloalkyl, —NR⁵R⁶, protected —OH, —CONR⁵R⁶, phosphonic acid,        sulfonic acid, phosphinic acid, —SO₂NR⁵R⁶, and a heterocyclic        methylene substituent as represented by Formula (III),

-   -   -   where        -   p is 0-6,        -   n is 0-2,        -   V, W, X and Z are each independently selected from O, S, and            NR¹⁶, where R¹⁶ is selected from: hydrogen, alkyl,            cycloalkyl, C₁-C₁₂aryl, substituted alkyl, substituted            cycloalkyl and substituted C₁-C₁₂aryl,        -   R⁴ is hydrogen, alkyl, cycloalkyl, C₁-C₁₂aryl, substituted            alkyl, substituted cycloalkyl and substituted C₁-C₁₂aryl,            and        -   R⁵ and R⁶ are each independently selected from hydrogen,            alkyl, substituted alkyl, C₃₋₆cycloalkyl, and aryl,        -   or R⁵ and R⁶ taken together with the nitrogen to which they            are attached represent a 5 to 6 member saturated ring            containing up to one other heteroatom selected from oxygen            and nitrogen;

    -   R¹⁵ is selected from the group consisting of alkyl, C₁-C₁₂aryl,        hydroxy, alkoxy, substituted alkyl, substituted C₁-C₁₂aryl and        halogen;

    -   m is 0-6; and

    -   Y is selected from alkyl, substituted alkyl and a cyclic or        polycyclic aromatic ring containing from 3 to 14 carbon atoms        and optionally containing from one to three heteroatoms,        provided that when the number of carbon atoms is 3 the aromatic        ring contains at least two heteroatoms and when the number of        carbon atoms is 4 the aromatic ring contains at least one        heteroatom, and optionally substituted with one or more        substituents selected from the group consisting of: alkyl,        substituted alkyl, C₁-C₁₂aryl, substituted cycloalkyl,        substituted C₁-C₁₂aryl, hydroxy, aryloxy, alkoxy, cycloalkyl,        nitro, cyano, halogen and protected —OH;

    -   and pharmaceutically acceptable salts, hydrates, solvates and        esters thereof;

    -   provided that at least one of R, R¹, R² and R³ is a substituted        aryl group or a heterocyclic methylene substituent as        represented in Formula (III).

Included among the presently invented compounds of Formula (II) arecompounds in which R¹⁵ is not alkoxy.

Included among the presently invented compounds of Formula (II) arethose having Formula (VI):

wherein:

-   -   R, R¹, R² and R³ are each independently selected from hydrogen,        C₁₋₆alkyl, C₁₋₆alkoxy, —(CH₂)_(p)OR⁴, —C(O)OR⁴, formyl, nitro,        cyano, halogen, aryl, substituted aryl, substituted alkyl,        —S(O)_(n)R⁴, cycloalkyl, —NR⁵R⁶, protected —OH, —CONR⁵R⁶,        phosphonic acid, sulfonic acid, phosphinic acid and —SO₂NR⁵R⁶,        -   where        -   p is 0-6,        -   n is 0-2,        -   R⁴ is hydrogen, alkyl, cycloalkyl, C₁-C₁₂aryl, substituted            alkyl, substituted cycloalkyl and substituted C₁-C₁₂aryl,            and        -   R⁵ and R⁶ are each independently selected from hydrogen,            alkyl, substituted alkyl, C₃₋₆cycloalkyl, and aryl,        -   or R⁵ and R⁶ taken together with the nitrogen to which they            are attached represent a 5 to 6 member saturated ring            containing up to one other heteroatom selected from oxygen            and nitrogen;    -   R¹⁵ is selected from the group consisting of alkyl, C₁-C₁₂aryl,        hydroxy, alkoxy, substituted alkyl, substituted C₁-C₁₂aryl and        halogen;    -   m is 0-6; and    -   Y is selected from alkyl, substituted alkyl and a cyclic or        polycyclic aromatic ring containing from 3 to 14 carbon atoms        and optionally containing from one to three heteroatoms,        provided that when the number of carbon atoms is 3 the aromatic        ring contains at least two heteroatoms and when the number of        carbon atoms is 4 the aromatic ring contains at least one        heteroatom, and optionally substituted with one or more        substituents selected from the group consisting of: alkyl,        substituted alkyl, C₁-C₁₂aryl, substituted cycloalkyl,        substituted C₁-C₁₂aryl, hydroxy, aryloxy, alkoxy, cycloalkyl,        nitro, cyano, halogen and protected —OH;    -   and pharmaceutically acceptable salts, hydrates, solvates and        esters thereof;    -   provided that at least one of R, R¹, R² and R³ is a substituted        aryl group.

Also included among the presently invented compounds of Formula (II) arecompounds of Formula (VI) in which R¹⁵ is not alkoxy.

Preferred among the presently invented Formula VI compounds are those inwhich, either:

-   -   R is a substituted aryl; and R¹ is hydrogen;        or:    -   R is hydrogen; and R¹ is a substituted aryl;        and in either case:    -   R² and R³ are each independently selected from hydrogen,        C₁₋₆alkyl, C₁₋₆alkoxy, nitro, cyano, halogen, aryl, substituted        aryl, substituted alkyl, cycloalkyl, phosphonic acid, phosphinic        acid and sulfonic acid;    -   R¹⁵ is selected from the group consisting of alkyl, substituted        alkyl, C₁-C₁₂aryl, alkoxy and halogen;    -   m is 0-4; and    -   Y is selected from,        -   phenyl, pyridinyl and pyrimidinyl, where the phenyl,            pyridinyl and pyrimidinyl are optionally substituted with            from one to three substituents selected from the group            consisting of: alkyl, substituted alkyl, C₁-C₁₂aryl,            substituted C₁-C₁₂aryl, alkoxy and halogen;    -   and pharmaceutically acceptable salts, hydrates, solvates and        esters thereof.

Particularly preferred among the presently invented Formula VI compoundsare those in which,

-   -   R is a substituted C₁-C₁₂aryl;        -   and    -   R¹ is hydrogen;    -   R² and R³ are each independently selected from hydrogen,        C₁₋₆alkyl, C₁₋₆alkoxy, nitro, cyano, halogen, substituted alkyl        and cycloalkyl;    -   R¹⁵ is selected from the group consisting of alkyl, substituted        alkyl, C₁-C₁₂aryl, alkoxy and halogen;    -   m is 0-2; and    -   Y is selected from,        -   phenyl, pyridinyl and pyrimidinyl, where the phenyl,            pyridinyl and pyrimidinyl are optionally substituted with            from one to three substituents selected from the group            consisting of: alkyl, substituted alkyl, C₁-C₁₂aryl,            substituted C₁-C₁₂aryl, alkoxy and halogen;    -   and pharmaceutically acceptable salts, hydrates, solvates and        esters thereof.

The most preferred among the presently invented Formula VI compounds arethose in which,

-   -   R is a substituted phenyl or pyridinyl ring; and    -   R¹ is hydrogen;    -   R² and R³ are each independently selected from hydrogen,        C₁₋₆alkyl, substituted alkyl and halogen;    -   R¹⁵ is selected from the group consisting of C₁₋₄alkyl,        C₁₋₄alkoxy, C₁-C₁₂aryl and halogen;    -   m is 0; and    -   Y is selected from,        -   phenyl, pyridinyl and pyrimidinyl, where the phenyl,            pyridinyl and pyrimidinyl is optionally substituted with            from one to three substituents selected from the group            consisting of: alkyl, substituted alkyl, C₁-C₁₂aryl,            substituted C₁-C₁₂aryl, alkoxy and halogen;    -   and pharmaceutically acceptable salts, hydrates, solvates and        esters thereof.

Preferred among the presently invented compounds are:

-   4′-{N′-[1-(3,4-Dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-3′-hydroxybiphenyl-4-carboxylic    acid;-   4′-{N′-[1-(3,4-Dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-3′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3,4-Dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(4-tert-Butylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   2-Aza-3′-{N′-[1-(4-tert-butylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-5′-chloro-2′-hydroxybiphenyl-3-carboxylic    acid;-   2-Aza-3′-{N′-[1-(4-tert-butylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3-Aza-3′-{N′-[1-(4-tert-butylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-5-carboxylic    acid;-   2-Aza-5′-chloro-3′-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   2-Aza-3′-{N′-[1-(4-tert-butylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxy-5′-methylbiphenyl-3-carboxylic    acid;-   2-Aza-3′-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxy-5′-methylbiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(4-tert-Butylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxy-5′-methylbiphenyl-3-carboxylic    acid;-   3-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-tetrazol-5-ylbiphenyl;-   3′-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-5′-fluoro-2′-hydroxybiphenyl-3-carboxylic    acid;-   7-({N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxyphenyl)quinolin-[1H]-one-3-carboxylic    acid;-   7-({N′-[1-(4-tert-butylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxyphenyl)quinolin-4[1H]-one-3-carboxylic    acid;-   3-Aza-3′-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-5-carboxylic    acid;-   3-Aza-3′-(N′-[1-{3-methyl-[4-(1-methylethyl)phenyl]-5-oxo-1,5-dihydropyrazol-4-ylidene}hydrazino)-2′-hydroxybiphenyl-5-carboxylic    acid;-   3-Aza-3′-{N′-[1-(4-tertbutylphenyl-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-5-carboxylic    acid;-   5′-Chloro-3′-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3,4-Dimethylphenyl)-3,5-dioxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(2-Ethoxy-2-oxoethyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-4′-(tetrazol-5-yl)biphenyl;-   3′-(N′-{1-[2-(N-tert-butyl)amino-2-oxoethyl]-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene}hydrazino)-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[3-Chloro-1-(3,4-dimethylphenyl)-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   5-chloro-3-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-4′-(tetrazol-5-yl)biphenyl;-   3′-{N′-[1-(3,4-Dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3,5-dicarboxylic    acid;-   3-Aza-3′-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxy-5′-methylbiphenyl-5-carboxylic    acid;-   3′-{N′-[1-(3,4-Dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-4-carboxylic    acid;-   3′-{N′-[1-(3,4-Dimethylphenyl)-3-methoxy-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(4-methoxyphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   (3-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-biphenyl)-1,1,1-trifluoromethanesulfonamide;-   3′-{N′-[1-(3,4-Dichlorophenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[3-methyl-5-oxo-1-(3-trifluoromethylphenyl)-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   8-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}quinolin-4[1H]-one-3-carboxylic    acid;-   3′-{N′-[3-methyl-5-oxo-1-(4-trifluoromethylphenyl)-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[3-methyl-5-oxo-1-(4-N-methylcarboxamidolphenyl)-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   N-[1-(3′-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-yl)methanoyl]methane    sulfonamide;-   3′-{N′-[3-methyl-5-oxo-1-phenyl-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[3-methyl-1-(4-methylphenyl)-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(4-chlorophenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(4-fluorophenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[3-methyl-5-oxo-1-(4-trifluoromethoxyphenyl)-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3,4-dimethylphenyl)-3-ethoxy-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3,4-dimethylphenyl)-3-(1-methylethoxy)-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[3-tert-butyl-1-(3,4-dimethylphenyl)-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[3-methyl-1-(4-methyl-2,3,5,6-tetrafluorophenyl)-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(4-fluoro-3-methylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3,4-dimethylphenyl)-3-phenyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3-{N′-[1-(3,4-dimethylphenyl)-5-oxo-3-phenyl-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-tetrazol-5-ylbiphenyl;-   3-{N′-[1-(3,4-dimethylphenyl)-3-methoxy-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-tetrazol-5-ylbiphenyl;-   3-{N′-[1-(3,4-dimethylphenyl)-3-ethoxy-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-tetrazol-5-ylbiphenyl;-   3-{N′-[1-(3,4-dimethylphenyl)-3-(1-methylethoxy)-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-tetrazol-5-ylbiphenyl;-   3-{N′-[1-(4-fluorophenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-tetrazol-5-ylbiphenyl;-   3-{N′-[1-(4-fluoro-3-methylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-tetrazol-5-ylbiphenyl;-   3-{N′-[3-methyl-5-oxo-1-(4-trifluoromethylphenyl)-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-tetrazol-5-ylbiphenyl;-   3′-{N′-[1-(3,4-dimethylphenyl)-3-(pyridin-4-yl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3-{N′-[1-(3,4-dimethylphenyl)-3-pyridin-4-yl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-tetrazol-5-ylbiphenyl;-   3-{N′-[1-(3,4-dimethylphenyl)-3-pyridin-2-yl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-tetrazol-5-ylbiphenyl;-   3′-{N′-[1-(3,4-dimethylphenyl)-3-(pyridin-2-yl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3-{N′-[1-(3-fluoro-4-methylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-tetrazol-5-ylbiphenyl;-   3′-{N′-[1-(3-fluoro-4-methylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[3-methyl-5-oxo-1-(4-trifluoromethylpyrimidin-2-yl)-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-N-tert-butoxycarbonylamino-3-{N′-[1-(3,4-Dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxybiphenyl;-   3′-amino-3-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxybiphenyl;-   3-{N′-[1-(3-fluorophenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-tetrazol-5-ylbiphenyl;-   3′-{N′-[1-(3-fluorophenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3-{N′-[3-methyl-5-oxo-1-(2,3,4,5,6-pentafluorophenyl)-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-tetrazol-5-ylbiphenyl;-   3′-{N′-[3-methyl-5-oxo-1-(2,3,4,5,6-pentafluorophenyl)-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3,4-difluorophenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3,4-dimethylphenyl)-3-methoxymethyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3-{N′-[1-(3,4-dimethylphenyl)-3-methoxymethyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-tetrazol-5-ylbiphenyl;-   3-{N′-[1-(3,4-difluorophenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-tetrazol-5-ylbiphenyl;-   3′-{N′-[1-(3,4-dimethylphenyl)-5-oxo-3-trifluoromethyl-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-6-fluoro-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3,4-dimethylphenyl)-5-oxo-3-propyl-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3-{N′-[1-(3,4-dimethylphenyl)-5-oxo-3-propyl-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-tetrazol-5-ylbiphenyl;-   3′-{N′-[1-(3,4-dimethylphenyl)-3-(1-methyl-1H-pyrrol-3-yl)-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3-{N′-[1-(3,4-dimethylphenyl)-3-(1-methyl-1H-pyrrol-3-yl)-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-tetrazol-5-ylbiphenyl;-   3′-{N′-[1-(3,4-dimethylphenyl)-3-furan-2-yl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3-{N′-[1-(3,4-dimethylphenyl)-3-furan-2-yl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-tetrazol-5-ylbiphenyl;-   N-(2′-hydroxy-3′-{N′-[3-methyl-5-oxo-1-(4-trifluoromethyl-phenyl)-1,5-dihydro-pyrazol-4-ylidene]hydrazino}biphenyl-3-yl)-1,1,1-trifluoromethanesulfonamide;-   N-(2′-hydroxy-3′-{N′-[1-(3-fluoro-4-methylphenyl)-3-methyl-5-oxo-1,5-dihydro-pyrazol-4-ylidene]hydrazino}biphenyl-3-yl)-1,1,1-trifluoromethanesulfonamide;-   N-(2′-hydroxy-3′-{N′-[1-(4-fluoro-3-methylphenyl)-3-methyl-5-oxo-1,5-dihydro-pyrazol-4-ylidene]hydrazino}biphenyl-3-yl)-1,1,1-trifluoromethanesulfonamide;-   N-(2′-hydroxy-3′-{N′-[1-(3,4-difluorophenyl)-3-methyl-5-oxo-1,5-dihydro-pyrazol-4-ylidene]hydrazino}biphenyl-3-yl)-1,1,1-trifluoromethanesulfonamide;-   N-(3′-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-yl)guanidine;-   3′-{N′-[1-(3,4-dimethylphenyl)-3-ethyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3-{N′-[1-(3,4-dimethylphenyl)-3-ethyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-tetrazol-5-ylbiphenyl;-   3′-{N′-[1-(3,4-dimethylphenyl)-5-oxo-3-thien-2-yl-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[3-cyclopropyl-1-(3,4-dimethylphenyl)-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3,4-dimethylphenyl)-5-oxo-3-thiazol-2-yl-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3,4-dimethylphenyl)-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3,4-dimethylphenyl)-3-(1-methylethyl)-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[3-(benzyloxymethyl)-1-(3,4-dimethylphenyl)-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[3-ethyl-5-oxo-1-(4-trifluoromethylphenyl)-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[5-oxo-1-(4-trifluoromethylphenyl)-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[-1-(3,4-dimethylphenyl)-3-hydroxymethyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[3-benzyloxymethyl-5-oxo-1-(4-trifluoromethylphenyl)-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[-1-(3,4-dimethylphenyl)-3-methylsulfanylmethyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[-1-(3,4-dimethylphenyl)-5-oxo-3-thiophen-3-yl-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[5-oxo-1-(4-trifluoromethylphenyl)-3-thiophen-3-yl-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[5-oxo-1-(4-trifluoromethylphenyl)-3-methylsulfanylmethyl-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   N-(3′-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydro-pyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-yl)methanesulfonamide;-   3′-[N′-(1-benzo[1,3]dioxol-5-yl-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene)hydrazino]-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3,5-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-4′-hydroxybiphenyl-4-carboxylic    acid;-   3′-{N′-[1-(3-chloro-4-methylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-4′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-phosphonic    acid;-   3′-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3,4-dicarboxylic    acid;-   2′,6-dihydroxy-3′-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}biphenyl-3-carboxylic    acid;-   4-aza-3′-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-5-carboxylic    acid;-   3′-{N′-[1-(3,4-dimethylphenyl)-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-sulfonic    acid; and-   5-(3′-{N′-[1-(3,4-Dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-ylmethylene)thiazolidine-2,4-dione;    and pharmaceutically acceptable salts, hydrates, solvates and esters    thereof.

Compounds of Formula (I) are included in the pharmaceutical compositionsof the invention and used in the methods of the invention.

By the term “protected hydroxy” or “protected —OH” as used herein, ismeant the alcoholic or carboxylic-OH groups which can be protected byconventional blocking groups in the art such as described in “ProtectiveGroups In Organic Synthesis” by Theodora W. Greene, Wiley-Interscience,1981, New York. Compounds containing protected hydroxy groups may alsobe useful as intermediates in the preparation of the pharmaceuticallyactive compounds of the invention.

By the term “aryl” as used herein, unless otherwise defined, is meant acyclic or polycyclic aromatic ring containing from 1 to 14 carbon atomsand optionally containing from one to five heteroatoms, provided thatwhen the number of carbon atoms is 1 the aromatic ring contains at leastfour heteroatoms, when the number of carbon atoms is 2 the aromatic ringcontains at least three heteroatoms, when the number of carbons is 3 thearomatic ring contains at least two heteroatoms and when the number ofcarbon atoms is 4 the aromatic ring contains at least one heteroatom.

By the term “C₁-C₁₂aryl” as used herein, unless otherwise defined, ismeant phenyl, naphthalene, 3,4-methylenedioxyphenyl, pyridine, biphenyl,quinoline, pyrimidine, quinazoline, thiophene, furan, pyrrole, pyrazole,imidazole and tetrazole.

When referring to compounds of Formula (I) and (II), the term“substituted” as used herein, unless otherwise defined, is meant thatthe subject chemical moiety has one or more substituents selected fromthe group consisting of: —CO₂R²⁰, aryl, —C(O)NHS(O)₂R²⁰, —NHS(O)₂R²⁰,hydroxyalkyl, alkoxy, —C(O)NR²¹R²², acyloxy, alkyl, amino, N-acylamino,hydroxy, —(CH₂)_(g)C(O)OR⁸, —S(O)_(n)R⁸, nitro, tetrazole, cyano, oxo,halogen, trifluoromethyl, protected —OH and a heterocyclic methylenesubstituent as represented by Formula (III),

where g is 0-6; R⁸ is hydrogen or alkyl; R²⁰ is selected form hydrogen,C₁-C₄alkyl, aryl and trifluoromethyl; R²¹ and R²² are independentlyselected from hydrogen, C₁-C₄alkyl, aryl and trifluoromethyl; V, W, Xand Z are each independently selected from O, S, and NR¹⁶, where R¹⁶ isselected from: hydrogen, alkyl, cycloalkyl, C₁-C₁₂aryl, substitutedalkyl, substituted cycloalkyl and substituted C₁-C₁₂aryl; and n is 0-2.

When referring to compounds of Formula (V) and (VI), the term“substituted” as used herein, unless otherwise defined, is meant thatthe subject chemical moiety has one or more substituents selected fromthe group consisting of: —CO₂R²⁰, aryl, —C(O)NHS(O)₂R²⁰, —NHS(O)₂R²⁰,hydroxyalkyl, alkoxy, —C(O)NR²¹R²², acyloxy, alkyl, amino, N-acylamino,hydroxy, —(CH₂)_(g)C(O)OR⁸, —S(O)_(n)R⁸, nitro, tetrazole, cyano, oxo,halogen, trifluoromethyl and protected —OH, where g is 0-6, R⁸ ishydrogen or alkyl, R²⁰ is selected form hydrogen, C₁-C₄alkyl, aryl andtrifluoromethyl, and R²¹ and R²² are independently selected formhydrogen, C₁-C₄alkyl, aryl and trifluoromethyl, and n is 0-2.

By the term “alkoxy” as used herein is meant —Oalkyl where alkyl is asdescribed herein including —OCH₃ and —OC(CH₃)₂CH₃.

The term “cycloalkyl” as used herein unless otherwise defined, is meanta nonaromatic, unsaturated or saturated, cyclic or polycyclic C₃-C₁₂.

Examples of cycloalkyl and substituted cycloalkyl substituents as usedherein include: cyclohexyl, 4-hydroxy-cyclohexyl, 2-ethylcyclohexyl,propyl 4-methoxycyclohexyl, 4-methoxycyclohexyl, 4-carboxycyclohexyl,cyclopropyl and cyclopentyl.

By the term “acyloxy” as used herein is meant —OC(O)alkyl where alkyl isas described herein. Examples of acyloxy substituents as used hereininclude: —OC(O)CH₃, —OC(O)CH(CH₃)₂ and —OC(O)(CH₂)₃CH₃.

By the term “N-acylamino” as used herein is meant —N(H)C(O)alkyl, wherealkyl is as described herein. Examples of N-acylamino substituents asused herein include: —N(H)C(O)CH₃, —N(H)C(O)CH(CH₃)₂ and—N(H)C(O)(CH₂)₃CH₃.

By the term “aryloxy” as used herein is meant —Oaryl where aryl isphenyl, naphthyl, 3,4-methylenedioxyphenyl, pyridyl or biphenyloptionally substituted with one or more substituents selected from thegroup consisting of: alkyl, hydroxyalkyl, alkoxy, trifluoromethyl,acyloxy, amino, N-acylamino, hydroxy, —(CH₂)_(g)C(O)OR⁸, —S(O)_(n)R⁸,nitro, cyano, halogen and protected —OH, where g is 0-6, R⁸ is hydrogenor alkyl, and n is 0-2. Examples of aryloxy substituents as used hereininclude: phenoxy, 4-fluorophenyloxy and biphenyloxy.

By the term “heteroatom” as used herein is meant oxygen, nitrogen orsulfur.

By the term “halogen” as used herein is meant a substituent selectedfrom bromide, iodide, chloride and fluoride.

By the term “alkyl” and derivatives thereof and in all carbon chains asused herein is meant a linear or branched, saturated or unsaturatedhydrocarbon chain, and unless otherwise defined, the carbon chain willcontain from 1 to 12 carbon atoms. Examples of alkyl substituents asused herein include: —CH₃, —CH₂—CH₃, —CH₂—CH₂—CH₃, —CH(CH₃)₂, —C(CH₃)₃,—(CH₂)₃—CH₃, —CH₂—CH(CH₃)₂, —CH(CH₃)—CH₂—CH₃, —CH═CH₂, and —C≡C—CH₃.

By the term “treating” and derivatives thereof as used herein, is meantprophylatic and therapeutic therapy.

All publications, including but not limited to patents and patentapplications, cited in this specification are herein incorporated byreference as though fully set forth.

Compounds of Formula (I) are included in the pharmaceutical compositionsof the invention and used in the methods of the invention. Where a —COOHor —OH group is present, pharmaceutically acceptable esters can beemployed, for example methyl, ethyl, pivaloyloxymethyl, and the like for—COOH, and acetate maleate and the like for —OH, and those esters knownin the art for modifying solubility or hydrolysis characteristics, foruse as sustained release or prodrug formulations.

The novel compounds of Formulas I and II are prepared as shown inSchemes I to IV below, or by analogous methods, wherein the ‘R’substituents, AR, Y and m are as defined in Formulas I and IIrespectively and provided that the ‘R’ and m substituents, AR and Y donot include any such substituents that render inoperative the processesof Schemes I to IV. All of the starting materials are commerciallyavailable or are readily made from commercially available startingmaterials by those of skill in the art.

i), nitric acid; sulfuric acid; ii) 4-carboxyphenylboronic acid;Pd(PPh₃)₄, Na2CO3, dioxane, water; iii) H₂, Pd—C; iv) NaNO₂, AR, NaHCO₃,water, EtOH

Scheme I outlines the formation of Formula I compounds. As used inscheme I, a 3-bromophenol (a) is nitrated with nitric acid or sodiumnitrate and sulfuric acid to give nitro phenol (b). Coupling of (b) witha substituted arylboronic acid, such as 3-carboxyphenylboronic acid or4-carboxyphenylboronic acid in the presence of a catalyst, preferablytetrakistriphenylphosphino palladium and a base such as sodium carbonateor triethylamine in a suitable solvent such as aqueous 1,4-dioxane ordimethylformamide afforded substituted aryl compound (c). Reduction ofthe nitro group by catalytic hydrogenation or mediated by a reducingmetal such as iron of tin dichloride in a suitable solvent such asethanol, acetic acid or water gives the aniline (d). Compound (d) isdiazotized by reaction with sodium nitrite and an appropriate acid, suchas nitric acid, sulfuric acid or, preferably, hydrochloric acid, in anappropriate aqueous solvent, such as water or, preferably anethanol-water mixture to produce a diazonium species which is directlyconverted to compound (e) in a coupling reaction with an appropriatearyl species in the presence of a base, preferably sodium hydrogencarbonate, or an acid, preferably hydrochloric acid.

i), NaNO₂, sulfuric acid; ii), MeI, K₂CO₃, acetone; iii)3-carboxyphenylboronic acid; Pd(PPh₃)₄, Na2CO3, dioxane, water; iv) 48%aqu. HBr, AcOH; v) H₂, Pd—C; yl) NaNO₂, AR, NaHCO₃, water, EtOH

Scheme II outlines an alternative synthesis of Formula I compounds. A2-bromophenol (f) (such as 2-bromophenol or 2-bromo-5-methylphenol isnitrated with nitric acid or sodium nitrate and sulfuric acid, to givenitro compound (g). The phenol (g) is then protected by reaction with analkylating agent such as benzyl bromide or preferably methyl iodide inthe presence of a base such as sodium hydride or potassium carbonate ina suitable solvent such as dimethylformamide, tetrahydrofuran or acetoneto give protected nitrophenol (h) (Prot=alkyl or substituted alkyl, e.g.methyl, benzyl). Coupling of (h) with a substituted arylboronic acid,such as 3-carboxyphenylboronic acid or 4-carboxyphenylboronic acid, inthe presence of a catalyst, preferably tetrakistriphenylphosphinopalladium and a base such as sodium carbonate to triethylamine in asuitable solvent such as aqueous 1,4-dioxane or dimethylformamideafforded substituted aryl compound (i). Removal of the protecting group(Prot) is accomplished using a protic or Lewis acid; such asconcentrated hydrobromic acid, boron tribromide or trimethylsilyl iodideto afford the phenol (j). Reduction of the nitro group by catalytichydrogenation or mediated by a reducing metal such as iron of tindichloride in a suitable solvent such as ethanol, acetic acid; or watergives the aniline (k). Compound (k) is diazotized by reaction withsodium nitrite and an appropriate acid, such as nitric acid, sulfuricacid or, preferably, hydrochloric acid, in an appropriate aqueoussolvent, such as water or, preferably, an ethanol-water mixture toproduce a diazonium species which is directly converted to compound (l)in a coupling reaction with an appropriate aryl species in the presenceof a base, preferably sodium hydrogen carbonate, or an acid, preferablyhydrochloric acid.

i) 5-(3-bromophenyl)tetrazole, Pd(PPh₃)₄, Na2CO3, dioxane, water; ii)48% aqu. HBr, AcOH; iii) HNO₂, AcOH; iv) H₂, Pd—C; v) NaNO₂, AR, NaHCO₃,water, EtOH

Scheme III outlines a further procedure for the synthesis of Formula Icompounds. A protected hydroxyphenylboronic acid; (m) (Prot=alkyl orsubstituted alkyl, e.g. methyl, benzyl) such as5-chloro-2-methoxyphenylboronic acid, 5-fluoro-2-methoxyphenyl, boronicacid or 2-methoxy-5-formylphenylboronic acid, is coupled with asubstituted halogenoaryl species, such as 5-(3-bromophenyl)tetrazole or5-bromonicotinic acid, in the presence of a catalyst, preferablytetrakistriphenylphosphino palladium, and a base, such as sodiumcarbonate or triethylamine in a suitable solvent such as aqueous1,4-dioxane or dimethylformamide afforded substituted aryl compound (n).Removal of the protecting group Prot is accomplished using an protic orLewis acid, such as concentrated hydrobromic acid, boron tribromide ortrimethylsilyl iodide to affored the phenol (o). Nitration of (o) withnitric acid, or sodium nitrate in the presence of an acid, such asacetic or hydrochloric acid, affords the nitro compound (p). Reductionof the nitro group by catalytic hydrogenation or mediated by a reducingmetal such as iron of tin dichloride in a suitable solvent such asethanol, acetic acid or water gives the aniline (q). Compound (q) isdiazotized by reaction with sodium nitrite and an appropriate acid, suchas nitric acid, sulfuric acid or, preferably, hydrochloric acid, in anappropriate aqueous solvent, such as water or, preferably, anethanol-water mixture to produce a diazonium species which is directlyconverted to compound (r) in a coupling reaction with an appropriatearyl species in the presence of a base, preferably sodium hydrogencarbonate, or an acid, preferably hydrochloric acid.

Scheme IV outlines the formation of pyrazoles for use in scheme I-III.An amine such as 4-methylaniline, compound (s), is diazotized by theaction of sodium nitrite and an appropriate acid, such as hydrochloricacid, nitric acid or sulfuric acid, in an appropriate aqueous solventsystem, such as water or ethanol-water mixtures, then reduced in situ bytin chloride to afford hydrazine, compound (t). The hydrazine is thencondensed with a electrophilic carbonyl species such as ethylacetoacetate (u), ethyl cyanoacetate or diethyl malonate, in anappropriate solvent such as acetic acid or ethanol at an appropriatetemperature typically 0-100° to give the corresponding pyrazole,compound (v) as described herein.

In preparing the presently invented compounds of Formula (I), thefollowing novel intermediates are prepared:

-   4′-Amino-3′-hydroxybiphenyl-4-carboxylic acid;-   4′-Amino-3′-hydroxybiphenyl-3-carboxylic acid;-   3′-Amino-2′-hydroxybiphenyl-3-carboxylic acid;-   3′-Amino-2′-hydroxybiphenyl-4-carboxylic acid;-   3-Amino-2-hydroxy-3′-(1H-tetrazol-5-yl)biphenyl;-   3-Amino-2-hydroxy-4′-(1H-tetrazol-5-yl)biphenyl;-   3-Amino-5-chloro-2-hydroxy-4′-(1H-tetrazol-5-yl)-biphenyl;-   6-(3-Amino-2-hydroxyphenyl)pyridine-2-carboxylic acid;-   6-(3-Amino-5-chloro-2-hydroxyphenyl)pyridine-2-carboxylic acid;-   6-(3-Amino-2-hydroxy-5-methylphenyl)pyridine-2-carboxylic acid;-   5-(3-Amino-2-hydroxyphenyl)nicotinic acid;-   5-(3-Amino-2-hydroxy-5-methylphenyl)nicotinic acid;-   2-(3-Amino-2-hydroxyphenyl)isonicotinic acid;-   3′-Amino-2′-hydroxy-5′-methylbiphenyl-3-carboxylic acid;-   3′-Amino-5′-fluoro-2′-hydroxybiphenyl-3-carboxylic acid;-   3′-Amino-5′-chloro-2′-hydroxybiphenyl-3-carboxylic acid;-   3′-Amino-2′-hydroxybiphenyl-3,5-dicarboxylic acid;-   N-[1-(3′-Amino-2′-hydroxybiphenyl-3-yl)methanoyl]methanesulfonamide;-   N-(3′-Amino-2′-hydroxybiphenyl-3-yl)-1,1,1-trifluoro-methanesulfonamide;-   (3′-Amino-2′-hydroxybiphenyl-3-yl)phosphonic acid;-   3′-Amino-2′-hydroxybiphenyl-3,4-dicarboxylic acid;-   3′-Amino-4,2′-dihydroxybiphenyl-3-carboxylic acid;-   3′-Amino-2′-hydroxybiphenyl-3-sulfonic acid;-   3′-Hydroxy-4′-nitrobiphenyl-4-carboxylic acid;-   3′-Hydroxy-4′-nitrobiphenyl-3-carboxylic acid;-   2′-Hydroxy-3′-nitrobiphenyl-3-carboxylic acid;-   2′-Hydroxy-3′-nitrobiphenyl-4-carboxylic acid;-   5-Chloro-2-hydroxy-3-nitro-3′-(1H-tetrazol-5-yl)biphenyl;-   5-Chloro-2-hydroxy-3-nitro-4′-(1H-tetrazol-5-yl)biphenyl;-   6-(5-Chloro-2-hydroxy-3-nitrophenyl)pyridine-2-carboxylic acid;-   6-(2-Hydroxy-5-methyl-3-nitrophenyl)pyridine-2-carboxylic acid;-   5-(5-Chloro-2-hydroxy-3-nitrophenyl)nicotinic acid;-   5-(5-Chloro-2-hydroxy-5-methyl-3-nitrophenyl)nicotinic acid;-   2-(5-Chloro-2-hydroxy-3-nitrophenyl)isonicotinic acid;-   5′-Chloro-2′-hydroxy-3′-nitrobiphenyl-3-carboxylic acid;-   5′-Chloro-2′-hydroxy-3′-nitrobiphenyl-3,5-dicarboxylic acid;-   N-[1-(5′-Chloro-2′-hydroxy-3′-nitrobiphenyl-3-yl)methanoyl]methanesulfonamide;-   1,1,1-Trifluoro-N-(2′-hydroxy-3′-nitrobiphenyl-3-yl)methanesulfonamide;-   (5′-Chloro-2′-hydroxy-3′-nitrobiphenyl-3-yl)phosphonic acid;-   5′-Chloro-2′-hydroxy-3′-nitrobiphenyl-3,4-dicarboxylic acid;-   5′-Chloro-4,2′-dihydroxy-3′-nitrobiphenyl-3-carboxylic acid;-   5′-Chloro-2′-hydroxy-3′-nitrobiphenyl-3-sulfonic acid;-   2′-Methoxy-3′-nitrobiphenyl-3-carboxylic acid;-   2′-Methoxy-3′-nitrobiphenyl-4-carboxylic acid;-   5-Chloro-2-hydroxy-3′-(1H-tetrazol-5-yl)biphenyl;-   5-Chloro-2-hydroxy-4′-(1H-tetrazol-5-yl)biphenyl;-   6-(5-Chloro-2-hydroxyphenyl)pyridine-2-carboxylic acid;-   6-(2-Hydroxy-5-methylphenyl)pyridine-2-carboxylic acid;-   6-(2-Hydroxy-5-methylphenyl)pyridine-2-carboxylic acid;-   5-(5-Chloro-2-hydroxy-5-methylphenyl)nicotinic acid;-   2-(5-Chloro-2-hydroxyphenyl)isonicotinic acid;-   5′-Chloro-2′-hydroxybiphenyl-3-carboxylic acid;-   5′-Chloro-2′-hydroxybiphenyl-3,5-dicarboxylic acid;-   N-[1-(5′-Chloro-2′-hydroxybiphenyl-3-yl)methanoyl]methanesulfonamide;-   3′-Amino-3-nitrobiphenyl-2-ol;-   (5′-Chloro-2′-hydroxybiphenyl-3-yl)phosphonic acid;-   5′-Chloro-2′-hydroxybiphenyl-3,4-dicarboxylic acid;-   5′-Chloro-4,2′-dihydroxybiphenyl-3-carboxylic acid;-   5′-Chloro-2′-hydroxybiphenyl-3-sulfonic acid;-   5-Chloro-2-methoxy-3′-(1H-tetrazol-5-yl)biphenyl;-   5-Chloro-2-methoxy-4′-(1H-tetrazol-5-yl)biphenyl;-   6-(5-Chloro-2-methoxyphenyl)pyridine-2-carboxylic acid;-   6-(2-Methoxy-5-methylphenyl)pyridine-2-carboxylic acid;-   6-(2-Methoxy-5-methylphenyl)pyridine-2-carboxylic acid;-   5-(5-Chloro-2-methoxy-5-methylphenyl)nicotinic acid;-   2-(5-Chloro-2-methoxyphenyl)isonicotinic acid;-   5′-Chloro-2′-methoxybiphenyl-3-carboxylic acid;-   5′-Chloro-2′-methoxybiphenyl-3,5-dicarboxylic acid;-   N-[1-(5′-Chloro-2′-methoxybiphenyl-3-yl)methanoyl]methanesulfonamide;-   N-(2′-Methoxy-3′-nitrobiphenyl-3-yl)-acetamide;-   (5′-Chloro-2′-methoxybiphenyl-3-yl)phosphonic acid;-   5′-Chloro-2′-methoxybiphenyl-3,4-dicarboxylic acid;-   5′-Chloro-4-hydroxy-2′-methoxybiphenyl-3-carboxylic acid; and-   5′-Chloro-2′-methoxybiphenyl-3-sulfonic acid.

The treatment of thrombocytopenia, as described herein, is accomplishedby increasing the production of platelets.

By the term “co-administering” and derivatives thereof as used herein ismeant either simultaneous administration or any manner of separatesequential administration of a TPO mimetic compound, as describedherein, and a further active ingredient or ingredients, known to treatthrombocytopenia, including chemotherapy-induced thrombocytopenia andbone marrow transplantation and other conditions with depressed plateletproduction. The term further active ingredient or ingredients, as usedherein, includes any compound or therapeutic agent known to or thatdemonstrates advantageous properties when administered with TPO or a TPOmimetic. Preferably, if the administration is not simultaneous, thecompounds are administered in a close time proximity to each other.Furthermore, it does not matter if the compounds are administered in thesame dosage form, e.g. one compound may be administered topically andanother compound may be administered orally.

Examples of a further active ingredient or ingredients for use incombination with the presently invented TPO mimetic compounds includebut are not limited to: chemoprotective or myeloprotective agents suchas G-CSF, BB 10010 (Clemons et al., Breast Cancer Res. Treatment, 1999,57, 127), amifostine (Ethyol) (Fetscher et al., Current Opinion inHemat., 2000, 7, 255-60), SCF, IL-11, MCP-4, IL-1-beta, AcSDKP (Gaudronet al., Stem Cells, 1999, 17, 100-6), TNF-a, TGF-b, MIP-1a (Egger etal., Bone Marrow Transpl., 1998, 22 (Suppl. 2), 34-35), and othermolecules identified as having anti-apoptotic, survival or proliferativeproperties.

Tpo has been demonstrated to act as a mobilizer of stem cells into theperipheral blood Neumann T. A. et al., Cytokines, Cell, & Mol. Ther.,2000, 6, 47-56). This activity can synergize with stem cell mobilizerssuch as G-CSF (Somolo et al., Blood, 1999, 93, 2798-2806). The TPOmimetic compounds of the present invention are thus useful in increasingthe numbers of stem cells in circulation in donors prior toleukapheresis for hematopoietic stem-cell transplantation in patientsreceiving myelo-ablative chemotherapy.

Likewise, TPO stimulates growth of myeloid cells, particularly those ofgranulocyte/macrophage lineage (Holly et al., U.S. Pat. No. 5,989,537).Granulocyte/macrophage progenitors are cells of the myeloid lineage thatmature as neutrophils, monocytes, basophils and eosinophils. Thecompounds described in the present invention have thus therapeuticutility in stimulating the proliferation of neutrophils in patients withneutropenic conditions.

Additional examples of a further active ingredient or ingredients foruse in combination with the presently invented TPO mimetic compoundsinclude but are not limited to: stem cell, megakaryocyte, neutrophilmobilizers such as chemotherapeutic agents (i.e., cytoxan, etoposide,cisplatin, Ballestrero A. et al., Oncology, 2000, 59, 7-13), chemokines,IL-8, Gro-beta (King, A. G. et al. J. Immun., 2000, 164, 3774-82),receptor agonist or antagonist antibodies, small molecule cytokine orreceptor agonists or antagonists, SCF, Flt3 ligand, adhesion moleculeinhibitors or antibodies such as: anti-VLA-4 (Kikuta T. et al., Exp.Hemat., 2000, 28, 311-7) or anti-CD44 (Vermeulen M. et al., Blood, 1998,92, 894-900), cytokine/chemokine/interleukin or receptor agonist orantagonist antibodies, MCP-4 (Berkhout T A., et al., J. Biol. Chem.,1997, 272, 16404-16413; Uguccioni M. et al., J. Exp. Med., 1996, 183,2379-2384).

Because the pharmaceutically active compounds of the present inventionare active as TPO mimetics they exhibit therapeutic utility in treatingthrombocytopenia and other conditions with depressed plateletproduction.

Likewise, the pharmaceutically active compounds of the present inventionare useful for enhancing platelet production obtained from a human donorprior to platelet pheresis, blood donation or platelet donation.

By the term “thrombocytopenia” and derivatives thereof as used herein isto be broadly interpreted as any decrease in the number of bloodplatelets below what is considered normal or desired for a healthyindividual. Thrombocytopenia is known to have many causative factors,including but not limited to, radiation therapy, chemotherapy, immunetherapy, immune thrombocytopenic purpura (ITP, Bussel J. B., Seminars inHematology, 2000, 37, Suppl 1, 1-49), myelodysplastic syndrom (MDS),aplastic anemia, AML, CML, viral infections (including, but not limitedto; HIV, hepatitis C, parvovirus) liver disease, myeloablation, bonemarrow transplant, stem cell transplant, peripheral blood stem celltransplant, progenitor cell defect, polymorphisms in stem cells andprogenitor cells, defects in Tpo, neutropenia (Sawai, N. J. LeukocyteBiol., 2000, 68, 137-43), dendritic cell mobilization (Kuter D. J.Seminars in Hematology, 2000, 37, Suppl 4, 41-49), proliferation,activation or differentiation. The pharmaceutically active compounds ofthis invention are useful in treating thrombocytopenia regardless of thefactor or factors causing the condition. The pharmaceutically activecompounds of this invention are also useful in treating thrombocytopeniawhen the causative factor or factors of the condition are unknown orhave yet to be identified.

Prophylactic use of the compounds of this invention is contemplatedwhenever a decrease in blood or blood platelets is anticipated.Prophylactic use of the compounds of this invention results in a buildup of platelets or a commencement of platelet production prior to ananticipated loss of blood or blood platelets. Prophylactic uses of thecompounds of this invention includes but is not limited to transplantsurgery, surgery, anesthesia prior to child birth and gut protection.

Human dendritic cells have been shown to express the TPO receptor(Kumamoto et al., Br. J. Haem, 1999, 105, 1025-1033) and TPO is a potentmobilizer of dendritic cells. The TPO mimetic compounds of the currentinvention are also useful as a vaccine adjuvant in that they increasethe activity and mobility of dendritic cells. The pharmaceuticallyactive compounds of this invention are useful as an immunologicaladjuvant, given in combination with an orally, transdermally orsubcutaneously delivered vaccine and/or immunomodulator, by increasingthe activity and mobility of dendritic cells.

Tpo is known to have various effects including anti-apototic/survivaleffects on megakaryocytes, platelets and stem cells, and proliferativeeffects on stem cells and megakaryocytic cells (Kuter D. J. Seminars inHematology, 2000, 37, 41-9). These Tpo activities effectively increasethe number of stem and progenitor cells so that there is synergisticeffects when Tpo is used in conjunction with other cytokines that inducedifferentiation.

The TPO mimetic compounds of the current invention are also useful inacting on cells for survival or proliferation in conjunction with otheragents known to act on cells for survival or proliferation. Such otheragents include but are not limited to: G-CSF, GM-CSF, TPO, M-CSF, EPO,Gro-beta, IL-11, SCF, FLT3 ligand, LIF, IL-3, IL-6, IL-1,Progenipoietin, NESP, SD-01, or IL-5 or a biologically active derivativeof any of the aforementioned agents, KT6352 (Shiotsu Y. et al., Exp.Hemat. 1998, 26, 1195-1201), uteroferrin (Laurenz J C., et al. Comp.Biochem. & Phys., Part A. Physiology., 1997, 116, 369-77), FK23(Hasegawa T., et al. Int. J. Immunopharm., 1996, 18 103-112) and othermolecules identified as having anti-apoptotic, survival or proliferativeproperties for stem cells, progenitor cells, or other cells expressingTpo Receptors.

In determining potency as TPO mimetics, the following assays wereemployed:

Luciferase Assay

Compounds of the present invention were tested for potency as mimeticsof the TPO receptor in a Luciferase assay such as described in Lamb, etal., Nucleic Acids Research 23: 3283-3289 (1995) and Seidel, et al.,Proc. Natl. Acad. Sci. USA 92: 3041-3045 (1995) by substituting aTPO-responsive BaF3 cell line (Vigon et al. Proc. Natl. Acad. Sci. USA1992, 89, 5640-5644) for the HepG2 cells utilized therein. The murineBaF3 cells express TPO receptors and closely match the pattern of STAT(signal transducers and activators of transcription) activation observedin primary murine and human bone marrow cells.

Proliferation Assay

Some of the more preferred compounds of this invention were active in anin vitro proliferation assay using the human UT7TPO cell line. UT7TPOcells are a human megakaryoblastic cell line that express Tpo-R, whosesurvival and growth is dependent on the presence of TPO (Komatsu et al.Blood 1996, 87, 4552).

Differentiation Assay

Likewise, some of the most preferred compounds of this invention werealso positive in stimulating the maturation of megakaryocytes from humanbone marrow cells. In this assay, purified human CD34+ progenitor cellswere incubated in liquid culture with test compounds for 10 days and thenumber of cells expressing the transmembrane glycoprotein CD41 (gpIIb),a megakaryocytic marker, was then measured by flow cytometry (seeCwirla, S. E. et al Science, 1997, 276, 1696).

The pharmaceutically active compounds within the scope of this inventionare useful as TPO mimetics in mammals, particularly humans, in needthereof.

Some of the preferred compounds within the scope of the invention showedactivation from about 4% to 100% control at a concentration of 0.001-10uM in the luciferase assay. The preferred compounds of the inventionalso promoted the proliferation of UT7TPO and 32D-mpI cells at aconcentration of 0.003 to 30 uM. The preferred compounds of theinvention also showed activity in the CD41 megakaryocytic assay at aconcentration of 0.003 to 30 uM.

The present invention therefore provides a method of treatingthrombocytopenia and other conditions with depressed plateletproduction, which comprises administering a compound of Formula (I) or apharmaceutically acceptable salt, hydrate, solvate or ester thereof in aquantity effective to enhance platelet production. The compounds ofFormula (I) also provide for a method of treating the above indicateddisease states because of their demonstrated ability to act as TPOmimetics. The drug may be administered to a patient in need thereof byany conventional route of administration, including, but not limited to,intravenous, intramuscular, oral, subcutaneous, intradermal, andparenteral.

The pharmaceutically active compounds of the present invention areincorporated into convenient dosage forms such as capsules, tablets, orinjectable preparations. Solid or liquid pharmaceutical carriers areemployed. Solid carriers include, starch, lactose, calcium sulfatedihydrate, terra alba, sucrose, talc, gelatin, agar, pectin, acacia,magnesium stearate, and stearic acid. Liquid carriers include syrup,peanut oil, olive oil, saline, and water. Similarly, the carrier ordiluent may include any prolonged release material, such as glycerylmonostearate or glyceryl distearate, alone or with a wax. The amount ofsolid carrier varies widely but, preferably, will be from about 25 mg toabout 1 g per dosage unit. When a liquid carrier is used, thepreparation will be in the form of a syrup, elixir, emulsion, softgelatin capsule, sterile injectable liquid such as an ampoule, or anaqueous or nonaqueous liquid suspension.

The pharmaceutical preparations are made following conventionaltechniques of a pharmaceutical chemist involving mixing, granulating,and compressing, when necessary, for tablet forms, or mixing, fillingand dissolving the ingredients, as appropriate, to give the desired oralor parenteral products.

Doses of the presently invented pharmaceutically active compounds in apharmaceutical dosage unit as described above will be an efficacious,nontoxic quantity preferably selected from the range of 0.001-100 mg/kgof active compound, preferably 0.001-50 mg/kg. When treating a humanpatient in need of a TPO mimetic, the selected dose is administeredpreferably from 1-6 times daily, orally or parenterally. Preferred formsof parenteral administration include topically, rectally, transdermally,by injection and continuously by infusion. Oral dosage units for humanadministration preferably contain from 0.05 to 3500 mg of activecompound. Oral administration, which uses lower dosages is preferred.Parenteral administration, at high dosages, however, also can be usedwhen safe and convenient for the patient.

Optimal dosages to be administered may be readily determined by thoseskilled in the art, and will vary with the particular TPO mimetic inuse, the strength of the preparation, the mode of administration, andthe advancement of the disease condition. Additional factors dependingon the particular patient being treated will result in a need to adjustdosages, including patient age, weight, diet, and time ofadministration.

The method of this invention of inducing TPO mimetic activity inmammals, including humans, comprises administering to a subject in needof such activity an effective TPO mimetic amount of a pharmaceuticallyactive compound of the present invention.

The invention also provides for the use of a compound of Formula (I) inthe manufacture of a medicament for use as a TPO mimetic.

The invention also provides for the use of a compound of Formula (I) inthe manufacture of a medicament for use in therapy.

The invention also provides for the use of a compound of Formula (I) inthe manufacture of a medicament for use in enhancing plateletproduction.

The invention also provides for the use of a compound of Formula (I) inthe manufacture of a medicament for use in treating thrombocytopenia.

The invention also provides for a pharmaceutical composition for use asa TPO mimetic which comprises a compound of Formula (I) and apharmaceutically acceptable carrier.

The invention also provides for a pharmaceutical composition for use inthe treatment of thrombocytopenia which comprises a compound of Formula(I) and a pharmaceutically acceptable carrier.

The invention also provides for a pharmaceutical composition for use inenhancing platelet production which comprises a compound of Formula (I)and a pharmaceutically acceptable carrier.

No unacceptable toxicological effects are expected when compounds of theinvention are administered in accordance with the present invention.

In addition, the pharmaceutically active compounds of the presentinvention can be co-administered with further active ingredients, suchas other compounds known to treat thrombocytopenia, includingchemotherapy-induced thrombocytopenia and bone marrow transplantationand other conditions with depressed platelet production, or compoundsknown to have utility when used in combination with a TPO mimetic.

Contemplated Equivalents—It will be appreciated by the person ofordinary skill in the art that the compounds of Formulas I and II mayalso exist in tautomeric forms. For example, in Formula I, the doublebond that is drawn between the two nitrogen atoms exists between thelower nitrogen atom and the AR substituent. Tautomeric forms of thecompounds of Formulas I and II are exemplified by the following Formula(IV):

where the ‘R’ groups are as defined above. All such compounds areincluded in the scope of the invention and inherently included in thedefinition of the compounds of Formulas I and II.

Without further elaboration, it is believed that one skilled in the artcan, using the preceding description, utilize the present invention toits fullest extent. The following Examples are, therefore, to beconstrued as merely illustrative and not a limitation of the scope ofthe present invention in any way.

EXPERIMENTAL DETAILS Example 1 Preparation of4′-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydroprazol-4-ylidene]hydrazino}-3′-hydroxybiphenyl-4-carboxylicAcid a) 5-bromo-2-nitrophenol

3-Bromophenol (32.9 g, 0.19 mol) was added slowly to a cold (10° C.)solution of sodium nitrate (29.0 g, 0.34 mol) in conc. sulfuric acid;(40.0 g) and water (70.0 mL) and the resulting mixture was allowed tostir at room temperature for 2 h. Water (200 mL) was added and theresulting mixture was extracted with diethyl ether and the extract wasdried (MgSO₄), filtered and concentrated. The residue was purified byflash chromatography (silica gel, 10% ethyl acetate/hexanes) to affordfirst the title compound (8.1 g, 20%), mp 40-42° C., then the undesiredisomer, 3-bromo-4-nitrophenol, as a yellow solid (12.7 g, 31%). mp125-127° C.

b) 3′-hydroxy-4′-nitrobiphenyl-4-carboxylic Acid

A solution of the compound from Example 1a) (2.18 g, 0.01 mol.),4-carboxyphenylboronic acid; (1.74 g, 0.0105 mol.), 2M aqu. sodiumcarbonate (10.0 mL; 0.02 mol.) and tetrakistriphenylphosphinopalladium(0) (0.5 g) in 1,4-dioxane (60.0 mL) was stirred and heatedunder reflux under a nitrogen atmosphere for 24 h.

The reaction mixture was cooled and evaporated and the residue treatedwith 6M aqu. hydrochloric acid; (100 mL). The grey precipitate wasfiltered and washed well with water then diethyl ether to afford thetitle compound (2.3 g; 88%) as a colorless solid. ¹H NMR (300 MHz,d₆-DMSO) δ 13.5-10.5 (br s, 2H), 8.06 (d, J=8.4 Hz, 2H), 8.03 (d, J=8.6Hz, 1H), 7.83 (d, J=8.4 Hz, 1H), 7.45 (d, J=1.8 Hz, 1H), 7.35 (dd,J=8.6, 1.8 Hz, 1H).

c) 4′-amino-3′-hydroxybiphenyl-4-carboxylic Acid; Hydrochloride Salt

A solution of the compound from Example 1b) (1.6 g, 0.0062 mol.) inethanol (75.0 mL), water (50.0 mL) and 3M aqu. sodium hydroxide (2.0 mL,0.0062 mol.) was hydrogenated over 10% palladium on carbon (0.2 g) atroom temperature and 50 psi for 2 h.

The reaction mixture was filtered, treated with 3M aqu. hydrochloricacid; (25.0 mL) then evaporated and the residue triturated with a littlewater to afford the title compound (1.18 g; 72%) as a brown solid. ¹HNMR (300 MHz, d₆-DMSO) δ 10.90 (s, 1H), 10.5-8.5 (br s, 3H), 8.03 (d,J=8.2 Hz, 2H), 7.71 (d, J=8.2 Hz, 2H), 7.41 (d, J=8.2 Hz, 1H), 7.36 (d,J=1.6 Hz, 1H), 7.22 (dd, J=8.2, 1.6 Hz, 1H).

d) 1-(3,4-Dimethylphenyl)-3-methyl-3-pyrazolin-5-one

A solution of 3,4-dimethylphenylhydrazine hydrochloride (17.7 g; 0.1mol.), ethyl acetoacetate (13.0 g; 0.1 mol.) and sodium acetate (8.2 g;0.1 mol.) in glacial acetic acid; (250 mL) was stirred and heated underreflux for 24 h.

The mixture was cooled and evaporated and the residue dissolved indiethyl ether (1 L) and carefully washed with sat. aqu. sodium hydrogencarbonate (5×200 mL). The ethereal layer was evaporated to afford thetitle compound (15.4 g; 76%). ¹H NMR (300 MHz, d₆-DMSO) δ 11.30 (br s,1H), 7.49 (d, J=1.4 Hz, 1H), 7.43 (dd, J=8.2 Hz, 1H), 7.14 (d, J=8.2 Hz,1H), 5.31 (s, 1H), 2.20 (s, 3H), 2.22 (s, 3H), 2.08 (s, 3H); MS (ES) m/z203 [M+H].

e)4′-{N′-[1-(3,4-Dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-3′-hydroxybiphenyl-4-carboxylicAcid; Hemihydrate

A suspension of the compound from Example 1c) (1.0 g; 0.0044 mol.) in 1Maqu. hydrochloric acid; (15.0 mL) was cooled to 5° C. then treateddropwise with a solution of sodium nitrite (0.32 g; 0.0046 mol.) inwater (5.0 mL). The yellow mixture was stirred at 5° C. for a further 10min. then treated in one portion with the compound from Example 1d)(0.882 g, 0.0044 mol.) followed by the portion-wise addition of sodiumhydrogen carbonate (1.8 g; 0.022 mol.) and ethanol (20.0 mL) ensuringthe final pH of the reaction mixture is approximately 7-8. The redsolution was then stirred at room temperature for 24 h.

The mixture was filtered to give a red solid which was slurried in water(50.0 mL) and then acidified with concentrated hydrochloric acid.Filtration afforded the title compound (0.68 g; 35%) as an orangepowder, mp=280° C. (dec.). ¹H NMR (300 MHz, d₆-DMSO) δ 13.62 (s, 1H),13.2-12.2 (br s, 1H), 10.92 (s, 1H), 8.02 (d, J=8.2 Hz, 2H), 7.73-7.69(m, 5H), 7.63 (d, 8.2 Hz, 1H), 7.31 (d, J=8.4 Hz, 1H), 7.29 (s, 1H),7.19 (d, J=8.4 Hz, 1H), 2.30 (s, 3H), 2.26 (s, 3H), 2.2 (s, 3H); Anal.(C₂₅H₂₂N₄O₄.0.5H₂O) calcd: C, 66.51; H, 5.13; N, 12.41. found: C, 66.74;H, 5.08; N, 12.36.

Example 2 Preparation of3′-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylicAcid a) 2-bromo-6-nitrophenol

Following the procedure of Example 1a) except substituting 2-bromophenolfor 3-bromophenol, the title compound was prepared (10.9 g; 25%) as abright, yellow solid. ¹H NMR (300 MHz, CDCl₃) δ 11.10 (S, 1 h), 8.13 (d,J=7.9 Hz, 1H), 7.89 (d, J=7.9 Hz, 1H), 6.90 (t, J=7.9 Hz, 1H).

b) 2-bromo-6-nitroanisole

a mixture of the compound from Example 2a) (10.8 g; 0.0495 mol.), methyliodide (3.4 mL; 0.00545 mol.) and potassium carbonate (8.2 g; 0.0592mol.) in acetone (250 mL) was stirred and heated under reflux for 24 h.

The mixture was evaporated and the residue triturated with water toafford the title compound (8.7 g; 76%). mp 55-56° C. ¹H NMR (300 MHz,CDCl₃ δ 7.81-7.74 (m, 2H), 7.13 (t, J=8.1 Hz, 1H), 4.02 (s, 3H); Anal.(C₇H₆NO₃Br) calcd: C, 36.24; H, 2.61; N, 6.04. found: C, 36.30; H, 2.59;N, 5.73.

c) 2′-methoxy-3′-nitrobiphenyl-3-carboxylic Acid

Following the procedure of Example 1b), except substituting the compoundfrom Example 2b) for 5-bromo-2-nitrophenol and substituting3-carboxyphenylboronic acid for 4-carboxyphenylboronic acid, the titlecompound was prepared (2.13 g; 47%) as a tan powder. ¹H NMR (300 MHz,d₆-DMSO) δ 8.12 (s, 1H), 8.03 (d, J=7.9 Hz, 1H), 7.94 (dd, J=7.9 Hz, 1.5Hz, 1H), 7.85 (d, J=7.9 Hz, 1H), 7.76 (dd, J=7.5, 1.5 Hz, 1H), 7.66 (t,J=7.5 Hz, 1H), 7.46 (t, j=7.9 Hz, 1H), 3.46 (s, 3H).

d) 2′-hydroxy-3′-nitrobiphenyl-3-carboxylic Acid

A solution of the compound from Example 2c) (2.13 g; 0.0077 mol.) inglacial acetic acid; (25.0 mL) and 48% aqu/hydrobromic acid; (25.0 mL)was stirred and heated under reflux for 5 h.

The mixture was cooled and filtered to afford the title compound (1.57g; 79%) as a tan powder. ¹H NMR (300 MHz, d₆-DMSO) δ 13.90 (s, 1H),10.66 (s, 1H), 8.12 (t, J=1.7 Hz, 1H), 8.07 (dd, J=8.4, 1.7 Hz, 1H),7.98 (dt, 7.8, 1.5 Hz, 1H), 7.79 (dt, J=8.1, 1.7 Hz, 1H), 7.74 (dd,J=7.5, 1.7 Hz, 1H), 7.62 (t, J=7.8 Hz, 1H), 7.17 (dd, J=8.4, 7.5 Hz,1H).

e) 3′-amino-2′-hydroxybiphenyl-3-carboxylic Acid; Hydrochloride Salt

Following the procedure of Example 1c), except substituting the compoundfrom Example 2d) for 3′-hydroxy-4′-nitrobiphenyl-4-carboxylic acid; thetitle compound was prepared (1.51 g; 100%) as a brown solid. 11.3-8.7(brs, 4H), 8.08 (s, 1H), 7.95 (d, J=7.8 Hz, 1H), 7.74 (d, J=7.8 Hz, 1H),7.61 (t, J=7.8 Hz, 1H), 7.34 (dd, J=7.8, 1.4 Hz, 1H), 7.24 (dd, J=7.8,1.3 Hz, 1H), 7.04 (t, J=7.8 Hz, 1H).

f)3′-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylicAcid; Hydrate

Following the procedure of Example 1e), except substituting the compoundfrom Example 2e) for 4′-amino-3′-hydroxybiphenyl-4-carboxylic acid;hydrochloride salt, the title compound was prepared (0.055 g; 32%) as anorange solid. mp 228° C. (dec.). ¹H NMR (300 MHz, d₆-DMSO) δ 13.76 (s,1H), 13.12 (s, 1H), 9.70 (s, 1H), 8.14 (s, 1H), 7.97 (dd, J=7.7 Hz, 1H),7.81 (dd, J=7.7 Hz, 1H), 7.74-7.60 (m, 5H), 7.22-7.13 (m, 3H), 2.34 (s,3H), 2.27 (s, 3H), 2.23 (s, 3H); Anal. (C₂₅H₂₂N₄O₄.1.0H₂O) calcd: C,65.21; H, 5.25; N, 12.17. found: C, 65.60; H, 4.96; N, 12.04.

Example 3 Preparation of3′-{N′-[1-(4-tert-butylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylicAcid; Hemihydrate a) 1-(4-tert-Butyl)-3-methyl-3-pyrazolin-5-one

Following the procedure of example 1d), except substituting4-tert-butylphenylhydrazine hydrochloride for3,4-dimethylphenylhydrazine hydrochloride, the title compound wasprepared (13.8 g; 60%). ¹H NMR (300 MHz, d₆-DMSO) δ 11.32 (s, 1H), 7.68(d, J=7.8 Hz, 2H), 7.40 (d, J=7.8 Hz, 2H), 5.32 (s, 1H), 2.09 (s, 3H),1.33 (s, 9H).

b)3′-{N′-[1-(4-tert-Butylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylicAcid; Hemihydrate

Following the procedure of Example 1e), except substituting the compoundfrom Example 2e) for 4′-amino-3′-hydroxybiphenyl-4-carboxylic acid;hydrochloride salt and the compound from Example 3a) for1-(3,4-dimethylphenyl)-3-methyl-3-pyrazolin-5-one, the title compoundwas prepared (0.391 g; 42%) as an orange solid, mp 145° C. (dec.). ¹HNMR (300 MHz, d₆-DMSO) δ 13.76 (s, 1H), 13.07 (s, 1H), 9.72 (s, 1H),8.14 (s, 1H), 7.98 (dd, J=7.8, 1.2 Hz, 1H), 7.83 (t, J=8.7 Hz, 1H), 7.73(dd, J=6.4, 3.1 Hz, 1H), 7.63 (t, J=7.7 Hz, 1H), 7.49 (d, J=7.8 Hz, 2H),7.20-7.16 (m, 2H), 2.35 (s, 3H), 1.31 (s, 9H). Anal. (C₂₇H₂₆N₄O₄.0.5H₂O)calcd: C, 67.63; H, 5.67; N, 11.68. found: C, 67.53; H, 5.46; N, 11.66.

Example 43-Aza-3′-{N′-[1-(4-tert-butylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-5-carboxylicAcid a) 5-(5-chloro-2-methoxyphenyl)-nicotinic Acid

Following the procedure of Example 1b), except substituting2-methoxy-5-chlorophenylboronic acid; for 4-carboxyphenylboronic acid;and substituting 5-bromonicotinic acid; for the compound of 1a), thetitle compound was prepared. MS (ES) m/z 264 [M+H].

b) 6-(5-chloro-2-hydroxyphenyl)-pyridine-2-carboxylic Acid

Following the procedure of Example 2d), except substituting the compoundof 4a) for the compound of 2c), the title compound was prepared. MS (ES)m/z 250 [M+H].

c) 6-(5-chloro-2-hydroxy-3-nitrophenyl)-pyridine-2-carboxylic Acid

To the solution of 6-(5-chloro-2-hydroxyphenyl)-pyridine-2-carboxylicacid; (2.3 g, 10.1 mmol) in 100 ml acetic acid; was added 1 ml fumingnitric acid; and stirred at 35° C. to 40° C. for half an hour. Thereaction mixture was diluted with water and adjusted pH to 2.5. Theresulting precipitation was collected, washed and dried to give solid(2.74 g; 78%, three steps). MS (ES) m/z 295 [M+H].

d)3-Aza-3′-{N′-[1-(4-tert-butylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-5-carboxylicAcid

Following the procedure of Example 1c), except substituting the compoundof 4c) for the compound of 1b), the crude product was isolated. Asuspension of the crude product (0.0015 mol.) in 1M aqu. hydrochloricacid; (25.0 mL) was cooled to 5° C. then treated dropwise with asolution of sodium nitrite (0.11 g; 0.0015 mol.) in water (5.0 mL). Theyellow mixture was stirred at 5° C. for a further 10 min. then treatedin one portion with the compound from Example 3a) (0.34 g, 0.0015 mol.)followed by the portion-wise addition of sodium hydrogen carbonate andethanol ensuring the final pH of the reaction mixture is approximately7-8. The red solution was then stirred at room temperature for 24 h. Themixture was filtered to give a red solid which was slurried in water(50.0 mL) and then acidified with concentrated hydrochloric acid.Filtration afforded the title compound (0.2 g; 29%) as a powder. ¹H NMR(300 MHz, d₆-DMSO) δ 13.8 (br, 2H), 9.9 (s, 1H), 9.08 (s, 1H), 8.9 (s,1H), 8.4 (s, 1H), 7.82 (d, J=7.7 Hz, 2H), 7.75 (d, J=7.9 Hz, 1H), 7.50(d, J=7.0 Hz, 2H), 7.20 (m, 2H), 2.34 (s, 3H), 1.32 (s, 9H) MS (ES) m/z472 (M+H)⁺.

Example 53-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-(tetrazol-5-yl)biphenyla) 5-(5′-chloro-2′-methoxybiphenyl-3-yl)-1H-tetrazole

Following the procedure of Example 1b), except substituting2-methoxy-5-chlorophenylboronic acid; for 4-carboxyphenylboronic acid;and substituting 5-(3-bromophenyl)-1H-tetrazole for the compound of 1a),the title compound was prepared (1.36 g; 100%) as a white solid. ¹H NMR(300 MHz, d₆-DMSO) δ 8.16 (s, 1H), 8.05 (d, J=7.6 Hz, 1H), 7.7 (d, J=6.6Hz, 1H), 7.67 (t, J=7.7 Hz, 1H), 7.48 (m, 2H), 7.2 (d, J=9.1 Hz, 1H),3.8 (s, 3H), MS (ES) m/z 287 [M+H].

b) 5-(5′-chloro-2′-hydroxybiphenyl-3-yl)-1H-tetrazole

Following the procedure of Example 2d), except substituting the compoundof 5a) for the compounds of 2c), the title compound was prepared. MS(ES) m/z 250 [M+H].

c) 5-(5′-chloro-2′-hydroxybiphenyl-3′-nitro-3-yl)-1H-tetrazole

Following the procedure of Example 4c), except substituting the compoundof 5b) for the compound of 4c), the title compound was prepared asyellow solid (0.5 g; 84%). ¹H NMR (300 MHz, d₆-DMSO) δ 8.2 (s, 1H), 8.1(d, J=5.9 Hz, 1H), 8.09 (d, J=7.4 Hz, 1H), 7.8 (d, J=2.7 Hz, 1H), 7.75(m, 2H), MS (ES) m/z 295 [M+H].

d)3-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-(tetrazol-5-yl)biphenyl

Following the procedure of Example 1c), except substituting the compoundof 5c) for the compound of 1b), the crude product was isolated. Asuspension of the crude product (0.0015 mol.) in 1M aqu. hydrochloricacid; (25.0 mL) was cooled to 5° C. then treated dropwise with asolution of sodium nitrite (0.11 g; 0.0015 mol.) in water (5.0 mL). Theyellow mixture was stirred at 5° C. for a further 10 min. then treatedin one portion with the compound from Example 1d) (0.34 g, 0.0015 mol.)followed by the portion-wise addition of sodium hydrogen carbonate andethanol ensuring the final pH of the reaction mixture is approximately7-8. The red solution was then stirred at room temperature for 24 h. Themixture was filtered to give a red solid which was slurried in water(50.0 mL) and then acidified with concentrated hydrochloric acid.Filtration afforded the title compound (0.14 g; 20%) as a powder. ¹H NMR(300 MHz, d₆-DMSO) δ 13.7 (s, 1H), 9.8 (s, 1H), 8.26 (s, 1H), 8.1 (d,J=1.5 Hz, 1H), 7.75 (m, 3H), 7.6 (d, J=2.2 Hz, 1H), 7.2 (m, 3H), 2.35(s, 3H), 2.25 (d, J=2.2 Hz, 6H).

Example 6 Capsule Composition

An oral dosage form for administering a presently invented agonist ofthe TPO receptor is produced by filling a standard two piece hardgelatin capsule with the ingredients in the proportions shown in TableI, below.

TABLE I INGREDIENTS AMOUNTS4′-{N′-[1-(3,4-Dimethylphenyl)-3-methyl-5-oxo-1,5- 25 mgdihydropyrazol-4-ylidene]hydrazino}-3′-hydroxybiphenyl-4- carboxylicacid; (Compound of Example 1) Lactose 55 mg Talc 16 mg MagnesiumStearate  4 mg

Example 7 Injectable Parenteral Composition

An injectable form for administering a presently invented agonist of theTPO receptor is produced by stirring 1.5% by weight of4′-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-3′-hydroxybiphenyl-3-carboxylicacid; in 10% by volume propylene glycol in water.

Example 8 Tablet Composition

The sucrose, calcium sulfate dihydrate and a presently invented agonistof the TPO receptor, as shown in Table II below, are mixed andgranulated in the proportions shown with a 10% gelatin solution. The wetgranules are screened, dried, mixed with the starch, talc and stearicacid; screened and compressed into a tablet.

TABLE II INGREDIENTS AMOUNTS3′-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5- 20 mgdihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3- carboxylicacid; (Compound of Example 2) calcium sulfate dihydrate 30 mg sucrose  4mg starch  2 mg talc  1 mg stearic acid 0.5 mg 

Preferred among the compounds of the present invention are thefollowing;

-   3′-{N′-[3-cyclopropyl-1-(3,4-dimethylphenyl)-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   [1-(4-fluoro-3-methylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[3-methyl-5-oxo-1-(4-trifluoromethylphenyl)-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3,4-Dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-tetrazol-5-ylbiphenyl;-   3′-{N′-[1-(3-fluoro-4-methylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3,4-dimethylphenyl)-3-ethyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3-{N′-[1-(3,4-dimethylphenyl)-3-ethyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-tetrazol-5-ylbiphenyl;-   3′-{N′-[1-(3-chloro-4-methylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-5′-fluoro-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3,4-Dimethylphenyl)-3-methoxy-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3-Aza-3′-{N′-[1-(4-tert-butylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-5-carboxylic    acid;-   3′-{N′-[3-methyl-1-(4-methylphenyl)-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   [1-(4-fluoro-3-methylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3,5-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   (3-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-biphenyl)-1,1,1,-trifluoromethanesulfonamide;    and-   3′-{N′-[1-(3,4-dimethylphenyl)-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid.

Particularly preferred among the compounds of the invention arefollowing;

-   3′-{N′-[3-cyclopropyl-1-(3,4-dimethylphenyl)-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   [1-(4-fluoro-3-methylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[3-methyl-5-oxo-1-(4-trifluoromethylphenyl)-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3,4-Dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-tetrazol-5-ylbiphenyl;-   3′-{N′-[1-(3-fluoro-4-methylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3,4-dimethylphenyl)-3-ethyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3-{N′-[1-(3,4-dimethylphenyl)-3-ethyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-tetrazol-5-ylbiphenyl;-   3′-{N′-[1-(3-chloro-4-methylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-5′-fluoro-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3,4-Dimethylphenyl)-3-methoxy-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3-Aza-3′-{N′-[1-(4-tert-butylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-5-carboxylic    acid; and-   3′-{N′-[3-methyl-1-(4-methylphenyl)-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid.

Particularly preferred among the compounds of the invention arefollowing;

-   3′-{N′-[3-cyclopropyl-1-(3,4-dimethylphenyl)-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   [1-(4-fluoro-3-methylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[3-methyl-5-oxo-1-(4-trifluoromethylphenyl)-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3-{N′-[1-(3,4-dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-tetrazol-5-ylbiphenyl;-   3′-{N′-[1-(3,4-Dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3-fluoro-4-methylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid;-   3′-{N′-[1-(3,4-dimethylphenyl)-3-ethyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylic    acid; and-   3-{N′-[1-(3,4-dimethylphenyl)-3-ethyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2-hydroxy-3′-tetrazol-5-ylbiphenyl.

The most preferred among the compounds of the invention is,3′-{N′-[1-(3,4-Dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylicacid.

The compound3′-{N′-[1-(3,4-Dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylicacid demonstrated an activity of, EC50=0.03 uM, 100% TPO in the aboveproliferation assay.

While the preferred embodiments of the invention are illustrated by theabove, it is to be understood that the invention is not limited to theprecise instructions herein disclosed and that the right to allmodifications coming within the scope of the following claims isreserved.

1. A method for enhancing platelet production in a human in need thereofwhich comprises administering to such human a therapeutically effectiveamount of a compound of the following formula:

wherein Q is —COOH; or a pharmaceutically acceptable salt thereof. 2.The method of claim 1 wherein the compound is administered orally. 3.The method of claim 1 wherein the compound is administered parenterally.4. The method of claim 1 further comprising co-administering atherapeutically effective amount of an agent selected from the groupconsisting of: a colony stimulating factor, cytokine, chemokine and aninterleukin or cytokine receptor agonist or antagonist.
 5. The method ofclaim 4 wherein the agent is selected from the group consisting of:G-CSF, GM-CSF, TPO, M-CSF, EPO, Gro-beta, IL-11, SCF, FLT3 ligand, LIE,IL-3, IL-6, IL-1, NESP, SD-01, IL-8 and IL-5.
 6. A method for enhancingplatelet production obtained from a human donor which comprisesadministering to such donor a therapeutically effective amount of thecompound3′-{N′-[1-(3,4-Dimethylphenyl)-3-methyl-5-oxo-1,5-dihydropyrazol-4-ylidene]hydrazino}-2′-hydroxybiphenyl-3-carboxylicacid or a pharmaceutically acceptable salt thereof; prior to plateletpheresis, blood donation or platelet donation.
 7. The method of claim 6wherein the compound is administered orally.
 8. The method of claim 6wherein the compound is administered parenterally.
 9. The method ofclaim 6 further comprising co-administering a therapeutically effectiveamount of an agent selected from the group consisting of: a colonystimulating factor, cytokine, chemokine and an interleukin or cytokinereceptor agonist or antagonist.
 10. The method of claim 9 wherein theagent is selected from the group consisting of: G-CSF, GM-CSF, TPO,M-CSF, EPO, Gro-beta, IL-11, SOF, FLT3 ligand, LIE, IL-3, IL-6, IL-1,NESP, SD-01, IL-8 and IL-5.